Mechanisms Underlying the Recovery of Lower Urinary Tract Function Following Spinal Cord Injury*

Home , Funiculus (neuroanatomy), Urinary bladder

Mechanisms underlying the recovery of lower urinary tract function following spinal cord injury*

WC de Groat Professor of Pharmacology and Neuroscience, University of Pittsburgh, School of Medicine, W1352 Biomedical Science Tower, Pittsburgh, PA 15261, USA

Keywords: neurourology; urinary bladder; neuroplasticity; automatic micturition; spinal cord injury

Introduction lower urinary tract: (1) a reservoir (the bladder) and The functions of the lower urinary tract to store and (2) an outlet (consisting of bladder neck, urethra, and periodically release urine are dependent upon neural striated muscles of the pelvic floor)_ Under normal circuits located in the brain, spinal cord and peripheral conditions, the urinary bladder and outlet exhibit a ganglia_1 ,2 This dependence on central nervous control reciprocal relationship. During urine storage, the distinguishes the lower urinary tract from many other bladder neck and proximal urethra are closed; and visceral structures (eg the gastrointestinal tract and the bladder smooth muscle is quiescent, allowing the cardiovascular system) that maintain a certain level of intravesical pressure to remain low over a wide range of function even after elimination of extrinsic neural bladder volumes. During voluntary micturition the input. initial event is a reduction of intraurethral pressure, The lower urinary tract is also unusual in regard to its which reflects a relaxation of the pelvic floor and the pattern of activity and the complexity of its neural paraurethral striated muscles and an opening of the regulation. For example, the urinary bladder has two bladder neck_4,5 These changes in the urethra are principal modes of operation: storage and elimination. followed in a few seconds by a detrusor contraction and Thus many of the neural circuits exhibit switch-like or a rise in intravesical pressure which is maintained until phasic patterns of activity3 in contrast to tonic patterns the bladder empties. Reflex inhibition of the smooth occurring in autonomic pathways to cardiovascular and striated muscles of the urethra also contribute to organs_ In addition, micturition is under voluntary the reduction of outlet resistance during micturition. control and depends upon learned behavior which These changes are coordinated by three sets of develops during maturation of the nervous system; nerves emerging from the thoracolumbar and sacral whereas many other visceral functions are regulated levels of the spinal cord: (1) sacral parasympathetic involuntarily_ Micturition also depends on the integra (pelvic nerves), (2) sacral somatic (pudendal nerves) tion of autonomic and somatic efferent mechanisms and (3) thoracolumbar sympathetic (hypogastric nerves within the lumbosacral spinal cord.4 This is necessary and sympathetic chain) (Figure 1).1-8 The sacral para during urine storage and elimination to coordinate the sympathetic efferent pathway provides the major exci activity of visceral organs (the bladder and urethra) tatory input to the bladder and consists of spinal with that of urethral striated muscles. preganglionic neurons which send axons to peripheral The dependence of lower urinary tract functions on ganglion cells which in turn innervate the bladder complex central neural networks renders these func smooth muscle (Figure 1)_ Transmission in bladder tions susceptible to a variety of neurological dis ganglia is mediated by nicotinic cholinergic receptors; orders_4,5 This paper will review studies in animals and whereas excitatory input to the bladder smooth muscle humans that have provided insights into the neural is mediated by acetylcholine acting on muscarinic control of the lower urinary tract and the disruption of receptors and by a cotransmitter adenosine triphos this control by spinal cord injury_ phate (ATP) acting on P2 purinergic receptors (Table 1)_9 Cholinergic transmission at both the ganglionic and Anatomy and innervation postganglionic level is modulated by a variety of The storage and periodic elimination of urine are transmitter mechanisms.2 For example ganglionic syn regulated by the activity of two functional units in the apses are regulated by muscarinic, adrenergic, puriner gic, and enkephalinergic mechanisms (Table 1)_ Post ganglionic transmission in the bladder is also controlled *The Sir Ludwig Guttmann Lecture given at the Annual Scientif presynaptically by transmitters which regulate acetyl Meeting of the International Medical Society of Paraplegia, Ghent, choline release_lO-13 The most prominent presynaptic Belgium, 27 May 1993 Spinal injury and micturition we de Groat

494 modulation is mediated by M-l muscarinic autorecep Lumbar tors which are activated during high frequency nerve spinal cord firing and markedly facilitate acetylcholine release.10,ll Thus, peripheral cholinergic synapses exhibit a con siderable degree of plasticity which is determined by Dorsal root the intensity and/or pattern of neural activity. This ganglia plasticity no doubt plays an important role in urine storage as well as in promoting complete emptying of the bladder. Pudendal N. Somatic efferent pathways to the external urethral sphincter are cholinergic and are carried in the pudendal nerve from anterior horn cells in the third and fourth sacral segments. Branches of the pudendal nerve and other sacral somatic nerves also carry efferent Urinary impulses to muscles of the pelvic floor. bladder Sympathetic preganglionic pathways that arise from the TIl to L2 spinal segments pass to the sympathetic chain ganglia and then to prevertebral ganglia in the superior hypogastric and pelvic plexuses and also to Diagram showing the sympathetic, parasympathe short adrenergic neurons in the bladder and urethra. tic,Figure and 1 somatic innervation of the urogenital tract of the male cat. Sympathetic preganglionic pathways emerge from Sympathetic postganglionic nerves which release nor the lumbar spinal cord and pass to the sympathetic chain epinephrine provide an excitatory input to smooth ganglia (SCG) and then via the inferior splanchnic nerves muscle of the urethra and bladder base, an inhibitory (ISN) to the inferior mesenteric ganglia (IMG). Pregan input to smooth muscle in the body of the bladder, as glionic and postganglionic sympathetic axons then travel in well as inhibitory and facilitatory input to vesical the hypogastric nerve (HGN) to the pelvic plexus and the parasympathetic ganglia.1,2,5 The adrenergic receptors urogenital organs. Parasympathetic preganglionic axons mediating these responses are listed in Table 1. which originate in the sacral spinal cord pass in the pelvic Other putative transmitters including neuropeptide nerve to ganglion cells in the pelvic plexus and to distal Y (NPY), vasoactive intestinal polypeptide (VIP) and ganglia in the organs. Sacral somatic pathways are con nitric oxide have also been identified in efferent tained in the pudendal nerve, which provides an innerva tion to the penis, the ischiocavernosus (IC), bulbocaver pathways to the lower urinary tract of animals and nosus (BC), and external urethral sphincter (EUS) muscles. humans (Table 1). NPY is present in adrenergic and The pudendal and pelvic nerves also receive postganglionic cholinergic neurons and when administered exogen axons from the caudal sympathetic chain ganglia. These ously has a prejunctional inhibitory action to suppress three sets of nerves contain afferent axons from the lumbo the release of norepinephrine and acetylcholine from sacral dorsal root ganglia. U: ureter; PG: prostate gland; postganglionic nerve terminals.12 Nitric oxide and VIP VD: vas deferens; Reproduced from de Groat, 19928 with have smooth muscle relaxant effects. Nitric oxide has permission been identified as the parasympathetic transmitter

Receptors for putative transmitters in the lower urinary tract Table 1 Tissue Cholinergic Adrenergic Other

Bladder body + (M2) - (f3z) + Purinergic (Pz) + (M3) + (lYI) - VIP + (M1)a + (lYI)a + Substance P - Neuropeptide ya Bladder base + (M2) + (lYI) - VIP + (M3) - Neuropeptide ya Ganglia + (N) + (lYj) - Enkephalinergic (D) + (M1) -(lYz) - Purinergic (Pj) + (f3) + Substance P Urethra + (M) + (lYj) + Purinergic (Pz) + (lYz) - VIP - (f3z) - Neuropeptide - Nitric oxide ya Sphincter striated muscle + (N)

Letters in parentheses indicate receptor type, M (muscarinic) and N (nicotinic) + and - indicate excitatory and inhibitory effects. aindicates presynaptic receptors on nerve terminals that control transmitter release Spinal injury and micturition we de Groat

495 mediating relaxation of the urethral smooth muscle.9,13 8 Bladder afferents Afferent activity arising in the bladder is conveyed to the central nervous system over both sets of autonomic nerves.1,5,14-19 The most important afferents for initiat ing micturition are those passing in the pelvic nerves to the sacral spinal cord. These afferents consist of small myelinated (AD) and unmyelinated (C) fibers which convey impulses from tension receptors and nocicep OCM tors in the bladder wall. Electrophysiological studies in the cat have shown that AD bladder afferents respond in a graded manner to passive distension as well as CC active contraction of the bladder.5 ,14 The intravesical o pressure threshold for activation of these afferents ranges from 5 to 15 mm Hg which is consistent with C-FOS pressures at which humans report the first sensation of b bladder filling during cystometry.

High threshold C-fiber afferents have also been '. detected in the cat bladder.15,16 Under normal condi \. . ' . .:','' "·I.� " tions the large majority of these afferents do not . respond to bladder distension and therefore have been .' OCM . : :":�'.,,::'.: "' .:.' ..•.. . .:: .... termed 'silent C-fibers'; however many of these fibers : .- : : can be activated by chemical irritation of the bladder �'." . mucosa15,16 or cold temperatures.20 Following chemical irritation, the C-fiber afferents exhibit spontaneous firing when the bladder is em ty and increased firing during bladder distension.15,1 p Activation of C-fiber afferents by chemical irritation facilitates the micturi Comparison of the distribution of bladder afferent tion reflex and decreases bladder capacity.1 ,17,21 Figureprojections 2 to the L6 spinal cord of the rat (A) with the c- os Administration of capsaicin, a neurotoxin which desen distribution of f positive cells in the L6 spinal segment following chemical irritation of the lower urinary tract of sitizes C-fiber afferents, blocks the bladder hyperactiv the rat (B). Afferents labelled by WGA-HRP injected into ity induced by noxious stimuli; but does not block the urinary bladder. C-fos immunoreactivity is present in micturition reflexes in normal animals indicating that the nuclei of cells. DH: dorsal horn; SPN: sacral parasym C-fiber afferent pathways are not essential for normal pathetic nucleus; CC: central canal. Calibration represents voiding. 1,17,22-24 500,um Immunohistochemical studies have shown that a large percentage of bladder afferent neurons contain 9 CNS peptides: calcitonin-gene-related-peptide (CG RP), Urinary vasoactive intestinal polypeptide (VIP) substance P, bladder \ Elimination Switch enkephalins and cholecystokinin (CCK) .1,18,25 In the \ spinal cord, peptidergic nerve terminals have a distri e--1 I�--!1 CJ bution very similar to the distribution of bladder Storage afferents (Figure 2) .19,26 In the bladder, these peptides are also common in nerves in the submucosal and subepithelial layers, and around blood vessels. The Urethral sphincter neurotoxin, capsaicin, when applied locally to the bladder in experimental animals releases peptides c from peripheral afferent terminals and produces in flammatory responses, including plasma extravasation Diagram illustrating the anatomy of the lower and vasodilation.17 These findings suggest that the Figureurinary 3 tract and the switchlike function of the micturition neuropeptides may be important transmitters in affer reflex pathway. During urine storage, a low level of affer ent pathways in the lower urinary tract. ent activity activates efferent input to the urethral sphincter. A high level of afferent activity induced by bladder distention activates the switching circuit in the central nervous system (CNS) , producing firing in the Reflex mechanisms controlling the lower urinary efferent pathways to the bladder, inhibition of the efferent tract outflow to the sphincter, and urine elimination The central pathways controlling lower urinary tract function are organized as simple on-off switching switching circuits are listed in Table 2 and are illus circuits (Figure 3) which maintain a reciprocal relation trated in Figure 4. Intravesical pressure measurements ship between the urinary bladder and the urethral during bladder filling in both man and animals reveal outlet. The principal reflex components of these low and relatively constant bladder pressures when the Spinal injury and micturition we de Groat

496 Reflexes to the lower urinary tract Table 2 Afferent pathway Efferent pathway Central pathway

Urine storage 1 External sphincter contraction (somatic nerves) Spinal reflexes Low level vesical afferent activity 2 Internal sphincter contraction (sympathetic nerves) (pelvic nerve) 3 Detrusor inhibition (sympathetic nerves) 4 Ganglionic inhibition (sympathetic nerves) 5 Sacral parasympathetic outflow inactive Micturition 1 Inhibition of external sphincter activity Spinobulbospinal reflexes High level vesical afferent activity 2 Inhibition of sympathetic outflow (pelvic nerve) 3 Activation of parasympathetic outflow

a 60 Infant bladder volume is below the threshold for inducing voiding (Figure 4A). The accommodation of the Reflex void bladder to increasing volumes of urine is primarily a o passive phenomenon dependent upon the intrinsic i Bladderfilling properties of the vesical smooth muscle and quiescence of the parasympathetic efferent pathway. In addi lA,S -- tion, in some species urine storage is also facilitated by o, 100 200 b sympathetic reflexes which mediate an inhibition of 60 Adult � Ql bladder activity, closure of the bladder neck and "0" Voluntary void Ql � contraction of the proximal urethra (Table 2, Figure 5). -g� 1 t--- --�=:=-: During bladder filling the activity of the sphincter iii� o - Start Stop: i Start electromyogram (EMG) also increases (Figure 4B), Bladderf ll ng i �... reflecting an increase in efferent firing in the pudendal . . .i .i . llll. �: �__ �_. .. __ nerve and an increase in outlet resistance which o 100 200 IIII300 �� contributes to the maintenance of urinary continence.

Paraplegic The storage phase of the urinary bladder can be switched to the voiding phase either involuntarily (reflexly) or voluntarily (Figure 4B). The former is readily demonstrated in the human infant (Figure 4A) Bladder�sphincter '------Bladderf ll g d or in the anesthetized animal when the volume of urine i i t exceeds the micturition threshold. At this point, .. • increased afferent firing from tension receptors in the o 100 200 300 .__ l. bladder reverses the pattern of efferent outflow, producing firing in the sacral parasympathetic pathways Combined cystometrograms and sphincter elec tromyogramsFigure 4 (EMG) comparing reflex voiding responses in and inhibition of sympathetic and somatic pathways. an infant (A) and in a paraplegic patient (C) with a The expulsion phase consists of an initial relaxation of voluntary voiding response in an adult (B). The abscissa in the urethral sphincter (Figure 4A) followed in a few all records represents bladder volume in milliliters and the seconds by a contraction of the bladder, an increase in ordinates represent bladder pressure in cm H20 and elec bladder pressure and flow of urine. Secondary reflexes trical activity of the EMG recording. On the left side of elicited by a flow of urine through the urethra facilitate each trace the arrows indicate the start of a slow infusion of bladder emptying.s,27 fluid into the bladder (bladder filling). Vertical dashed lines These reflexes require the integrative action of indicate the start of sphincter relaxation which precedes by neuronal populations at various levels of the neuraxis a few seconds the bladder contraction in A and B. In part (Figures 5 and 6). Certain reflexes, for example, those B note that a voluntary cessation of voiding (stop) is associated with an initial increase in sphincter EMG fol mediating the excitatory outflow to the sphincters and lowed by a reciprocal relaxation of the bladder. A resump the sympathetic inhibitory outflow to the bladder, are tion of voiding is again associated with sphincter relaxation organized at the spinal level (Figure 5), whereas the and a delayed increase in bladder pressure. On the other parasympathetic outflow to the detrusor has a more hand, in the paraplegic patient (C) the reciprocal relation complicated central organization involving spinal and ship between bladder and sphincter is abolished. During spinobulbospinal pathways (Figure 6). bladder filling, transient uninhibited bladder contractions occur in association with sphincter activity. Further filling leads to more prolonged and simultaneous contractions of the bladder and sphincter (bladder-sphincter dyssynergia). Anatomy of spinal reflex circuits Loss of the reciprocal relationship between bladder and The spinal circuitry controlling the lower urinary tract sphincter in paraplegic patients interferes with bladder consists of three basic components: efferent neurons, emptying. Reproduced from de Groat and Steers, 19907 interneurons and primary afferent neurons. New with permission research methodology including transneuronal virus Spinal injury and micturition we d. Groat

497 Sphincter reflexes Cortical diencephalic mechanism

Micturition center

Inhibits sympathetic and sphincter x reflexes

Capsaicin block S�i� a�:�� t ,/ Lumbar cord c 4---- -L9 Unmyelinated__ affere___nt s � Ves ical IC) '------[ affere nts L Ganglia [r-- ---I _D e_r_ut s_o_r - Hypogastric L- --l nerve Diagram of the central reflex pathways that regu Contracts I lateFigure micturition 6 in the cat. In an animal with an intact bladder outlet I I neuraxis, micturition is initiated by a supraspinal reflex Inhibits ± : pathway passing through a center in the brainstem. The d pathway is triggered by myelinated afferents (Ab) con nected to tension receptors in the bladder wall. In spinal � >:: animals, connections between the brainstem and the sacral Urinary Sacral cord Pelvic nerve () !)..� spinal cord are interrupted (X) and micturition is initially bladder � �� blocked. In chronic spinal animals, however a spinal reflex mechanism emerges which is triggered by unmyelinated (C-fiber) vesical afferents. The C-fiber reflex pathway is + Pudendal nerve usually weak or undetectable in animals with an intact

External nervous system. Capsaicin (20-30 mg Kg-l s.c.) blocks the sphincter C-fiber reflex in chronic cats, but does not block micturi tion reflexes in intact cats. Reproduced from de Groat et Diagram showing detrusor-sphincter reflexes. at, 199023 with permission DuringFigure 5 the storage of urine, distention of the bladder produces low level vesical afferent firing, which in turn stimulates (1) the sympathetic outflow to the bladder outlet An unusual feature of sacral preganglionic neurons (base and urethra) and (2) pudendal outflow to the external in the cat is an extensive axon collateral system which urethral sphincter. These responses occur by spinal reflex projects bilaterally to various regions of the dorsal and pathways and represent 'guarding reflexes', which promote ventral horns, including: the area around the central continence. Sympathetic firing also inhibits detrusor muscle canal; the intermediolateral region of the sacral para and transmission in bladder ganglia. At the initiation of sympathetic nucleus, the dorsal commissure and lateral micturition, intense vesical afferent activity activates the dorsal horn.29 It has been speculated that these brainstem micturition center, which inhibits the spinal guarding reflexes collaterals mediate intraspinal integrative actions of preganglionic neurons including recurrent inhibition and modulation of sphincter reflexes. tracing, intracellular labelling, measurements of gene Sphincter motoneurons expression and patch clamp recording in spinal cord Retrograde axonal tracing has identified pudendal slice preparations have recently provided many new motoneurons and a subpopulation of these neurons insights into the morphological and electrophysio innervating the external urethral sphincter logical properties of these reflex components. (EUS) .1,5.30-32 In the cat EUS motoneurons are located in the ventrolateral division of Onut's nucleus and send Preganglionic neurons dendritic projections into: (1) the lateral funiculus, Preganglionic neurons in the lumbosacral parasympa (2) lamina X, (3) intermediolateral grey matter and thetic nucleus send dendrites to discrete regions of the (4) rostrocaudally within the nucleus (Figure 7). The spinal cord including: (1) the lateral and dorsal lateral dentrites occur in bundles passing through restricted funiculus, (2) lamina I on the lateral edge of the dorsal regions of the spinal cord. Thus the dendritic distribu horn, (3) the dorsal grey commissure and (4) grey tion of sphincter motoneurons (ie lateral, dorsolateral matter and lateral funiculus ventral to the autonomic and dorsomedial) is similar to that of sacral pregangli nucleus.1•28 As shown in Figure 7 the cells have long, onic neurons indicating that these two populations of relatively unbranched dendrites that can extend for neurons may receive synaptic inputs from the same distances of 1.5-2 mm in both the grey and white interneuronal pathways and fiber tracts in the spinal matter. 28 cord. Spinal injury and micturition We d. Groat

498 dorsal horn and then pass rostrocaudally gIVIng off collaterals that extend laterally and medially through the superficial layer of the dorsal horn (lamina I) into the deeper layers (laminae V-VII and X) at the base of the dorsal horn (Figure 2A). The lateral pathway which is the most prominent projection terminates in the region of the sacral parasympathetic nucleus (SPN) and also sends some axons to the dorsal commissure (Figure 2A). Bladder afferents have not been detected in the center of the dorsal horn (laminae III -IV) or in the ventral horn. The spinal neurons involved in processing afferent input from the lower urinary tract have been identified by the expression of the immediate early gene, c-fos (Figure 2B).21.38 The protein product of the c-fos gene can be detected immunocytochemically in the nuclei of neurons within 30-60 min following synaptic activa Comparison of the dendritic distributions of exter tion. In the rat noxious21 or non-noxious38 stimulation nalFigure urethral 7 sphincter motoneurons (left side) and a sacral parasympathetic preganglionic neuron (right side) in the of the bladder and urethra as well as electrical cat. The sacral preganglionic neuron which was filled intra stimulation of the pelvic nerves increased the levels of cellularly with neurobiotin exhibits dendrites which extend fos protein primarily in the dorsal commissure and in into: (1) the lateral funiculus, (2) lateral lamina I, (3) the the area of the sacral parasympathetic nucleus, the dorsal commissure and (4) within the sacral autonomic major sites of termination of afferents from the lower nucleus. Dendrites were reconstructed from serial sections urinary tract (Figure 2A). extending 500 11m rostral and caudal to the cell body. Two sphincter motoneurons (left side) were labelled by retro grade axonal transport following injections of cholera toxin HRP into the external urethral sphincter. Although the Storage reflexes dendritic distributions are incomplete, they show prominent Sympathetic pathways dendritic projections of one cell to lamina X around the Studies in animals indicate that sympathetic input to central canal and projections of another cell to the interme the lower urinary tract is tonically active during bladder diolateral region. Right and left sides are at slightly differ filling.1,3 Surgical or pharmacologic blockade of the ent rostrocaudal levels. Calibration represents 500 11m sympathetic pathways can reduce urethral resistance, reduce bladder capacity, reduce bladder wall com pliance and increase the frequency and amplitude of Interneurons bladder contractions.1 Tract tracing using viruses which cross synapses has Sympathetic firing is initiated at least in part by a been very effective in identifying spinal interneuronal sacrolumbar intersegmental spinal reflex pathway as well as brain pathways involved in the control of the which is triggered by vesical afferent activity in the lower urinary tract.33-37 As shown in Figure 8, inter pelvic nerves (Figure 5).1.3 This vesicosympathetic neurons retrogradely labelled by injection of pseudora reflex represents a negative feedback mechanism bies virus into the urinary bladder of the rat are located whereby an increase in bladder pressure triggers an in restricted regions of the spinal cord including the increase in inhibitory input to the bladder thus allowing area surrounding the autonomic nucleus, the dorsal the bladder to accommodate larger volumes of urine. commissure and the superficial laminae of the dorsal horn. Interneuronal locations overlap in many respects with the dendritic distribution of the efferent neurons. Somatic efferent pathways to the urethral sphincter A similar distribution of labelled interneurons has been The reflex control of the striated urethral sphincter is noted following injections of virus into the urethra35 or similar to the control of the sympathetic outflow to the 7 the external urethral sphincter3 indicating a prominent lower urinary tract (Figure 5). During bladder filling overlap of the interneuronal pathways controlling the pudendal motoneurons are activated by vesical afferent various target organs of the lower urinary tract. input, whereas during micturition the motoneurons are reciprocally inhibited. The inhibition is dependent in part on supraspinal mechanisms since in chronic spinal Afferent projections in the spinal cord animals and in paraplegic patients it is weak or absent. Afferent pathways from the LUT project to discrete In spinal injured patients the uninhibited spinal vesico regions of the dorsal horn that contain the interneurons sphincter excitatory reflex pathway commonly initiates as well as the soma and/or dendrites of efferent a striated sphincter contraction in concert with a neurons innervating the LUT. Pelvic nerve afferent contraction of the bladder (bladder-sphincter dyssyn pathways from the urinary bladder of the cat1.19 and ergia) (Figure 4C).4-6 This reflex interferes with blad rat26 project into Lissauer's tract at the apex of the der emptying. Spinal injury and micturition we de Groat

499 Voiding reflexes neurons.49 Thus, glutamate is likely to be involved at Micturition is mediated by activation of the sacral various sites in the micturition reflex pathway. parasympathetic efferent pathway to the bladder and reciprocal inhibition of the somatic pathway to the Pontine micturition center (PMC) urethral sphincter (Table 2; Figure 4B). Studies in cats Physiological and anatomical experiments have pro using brain lesioning techniques revealed that neurons vided substantial support for the concept that neuronal in the brainstem at the level of the inferior colliculus circuitry in the PMC functions as a switch in the have an essential role in the control of the parasympa micturition reflex pathway. The switch seems to regu thetic component of micturition (Figure 6). Removal of late bladder capacity and also coordinates the activity areas of the brain above the inferior colliculus by of the bladder and of the external urethral sphincter.1 ,39 intercollicular decerebration usually facilitates micturi Electrical or chemical stimulation in the PMC of the tion by elimination of inhibitory inputs from more rat, cat and dog induces: (1) a suppression of urethral rostral centers.l.S·27 However, transections at any point sphincter EMG, (2) firing of sacral preganglionic below the colliculi abolish micturition. Bilateral lesions neurons, (3) bladder contractions and (4) release of in the rostral pons in the region of the locus coeruleus urine. 1.3,5.50 On the other hand, microinjections of in cats or the lateral dorsal tegmental nucleus in rats putative inhibitory transmitters into the PMC of the cat abolishes micturition, whereas electrical or chemical can increase the volume threshold for inducing micturi stimulation at these sites triggers bladder contractions tion and in high doses completely block reflex voiding and micturition.1.3.2 7.39-41 These observations led to indicating that synapses in this region are important for the concept of a spinobulbospinal micturition reflex regulating the set point for reflex voiding and also are pathway that passes through a center in the rostral an essential link in the reflex pathway. 39 In addition brainstem (the pontine micturition center, PMC) administration of antagonists for certain inhibitory (Figure 6). The pathway functions as 'on-off' switch transmitters (eg the GABA-antagonist, bicuculline or (Figure 3) which is activated by a critical level of the opioid antagonist, naloxone) can reduce the mic afferent activity arising from tension receptors in the turition volume threshold indicating that the setpoint is bladder and is in turn modulated by inhibitory and under tonic inhibitory modulation. excitatory influences from areas of the brain rostral Virus tracing studies in the rat have revealed to the pons (eg diencephalon and cerebral cortex) extensive labelling in the PMC (Figure 8) following (Figure 6). injections of virus into the bladder, urethra and the external urethral sphincter.33 -37 These data are consist ent with the view that neurons in this region coordinate Spinobulbospinal micturition reflex pathway the activity of all the major organs involved in voiding. Electrophysiological studies in cats and rats have confirmed that the parasympathetic efferent outflow to the urinary bladder is activated by a long latency supraspinal reflex pathway.1.3.42-44 In cats recordings Recovery of lower urinary tract function from sacral parasympathetic preganglionic neurons following spinal cord injury innervating the urinary bladder show that reflex firing Spinal cord injury rostral to the lumbosacral level occurs with a long latency (65-100 ms) following eliminates voluntary and supraspinal control of void stimulation of myelinated (AD) vesical afferents in the ing, leading initially to an areflexic bladder and pelvic nerve.1.3.44 Afferent stimulation also evokes complete urinary retention followed by a slow develop negative field potentials in the rostral pons at latencies ment of automatic micturition and bladder hyperactiv of 30-40 ms; whereas electrical stimulation in the pons ity mediated by spinal pathways.l.4-6,44,51-53 However, excites sacral preganglionic neurons at latencies of voiding is commonly inefficient due to simultaneous 45-60 ms. The sum of the latencies for the spinobulbar contractions of the bladder and the urethral sphincter and bulbospinal components of the reflex pathway (bladder-sphincter dyssynergia). approximate the latency for the entire reflex. Recent experiments have also provided information regarding the transmitter in the descending limb of the Reorganization of reflex pathways micturition reflex pathway.1 Glutamic acid has at Electrophysiological studies in rats and cats have tracted most interest since it is the major excitatory shown that the reflex pathways in intact and chronic transmitter in the central nervous system. Experiments animals are markedly different. In both species, the in rats indicate that glutamatergic transmission in the central delay for the micturition reflex in chronic spinal cord is essential for bladder and urethral spinal animals is considerably shorter ( 5 ms in sphincter reflexes and for the spinal processing of rats; 15-40 ms in cats) than in intact< animals afferent input from the urinary bladder.45- 48 Both (60-75 ms). 1,3.42.44 In addition, in chronic spinal cats the NMDA and non-NMDA glutamatergic receptors have afferent limb of the micturition reflex consists of been implicated in transmission between the brain unmyelinated (C-fiber) afferents, whereas in intact cats micturition center and the spinal preganglionic it consists of myelinated (AD) afferents (Figure 6).23 neurons. These receptors also mediate excitatory trans This was demonstrated with electrophysiological mission between interneurons and preganglionic recording and also by administering capsaicin,23 a Spinal injury and micturition Wed. Groat

500

Cerebral cort ex

Pont in e micturit ion cent er Hypot halamus I PVN MPOA PeriV.N. nuclei AS LC I .. ". � ' IF DCM

Virus

�

Transneuronal virus tracing of the central pathways controlling the urinary bladder of the rat. Injection of pseudorabiesFigure 8 virus into the wall of the urinary bladder leads to retrograde transport of virus and sequential infection of postganglionic neurons, preganglionic neurons and then various central neural circuits synaptically linked to the pregangli onic neurons. At long survival times virus can be detected with immunocytochemical techniques in neurons at specific sites throughout the spinal cord and brain extending to the pontine micturition center in the pons (ie Barrington's nucleus or the laterodorsal tegmental nucleus) and to the cerebral cortex. Other sites in the brain labeled by virus are: (1) the paraventricular nucleus (PVN), medial preoptic area (MPOA) and periventricular nucleus (Peri V.N.) of the hypothalamus, (2) periaqueductal gray (PAG), (3) locus coeruleus (LC) and subcoeruleus, (4) red nucleus (RN), (5) medullary raphe nucleus and (6) the noradrenergic cell group designated AS. L6 spinal cord section showing on the left side the distribution of virus labelled parasympathetic preganglionic neurons (D) and interneurons (e) in the region of the parasympathetic nucleus 72 h after injection of the virus into the bladder. Interneurons in the dorsal commissure and the superficial laminae of the dorsal horn are not shown. The left side shows the entire population of preganglionic neurons (PGN) labeled by axonal tracing with the fluorescent dye fluorogold, injected into the pelvic ganglia. The right side shows the distribution of virus-labeled bladder PGN (_) among the entire population of PGN (D). Composite diagram of neurons in 12 spinal sections (42.um)

neurotoxin which is known to disrupt the function of bladder distension (ie silent C-fibers)Y However, in C-fiber afferents.17 In normal cats, capsaicin injected chronic spinal cats bladder distension initiates auto systemically in large doses (30-45 mg, s.c.) did not matic micturition by activating C-fiber afferent block reflex contractions of the bladder or the AD-fiber neurons.23 Thus, spinal injury must change the proper evoked bladder reflex. However in chronic spinal cats ties of these afferent receptors in the bladder. (3-6 weeks after spinal transection) capsaicin Other reflexes that appear following spinal cord (20-30 mgjkg, s.c.) completely blocked the rhythmic injury also may be mediated by C-fiber afferents. For bladder contractions induced by bladder distension and example, it is known that instillation of cold water into blocked the C-fiber-evoked reflex firing recorded on the bladder of patients with upper motoneuron lesions bladder postganglionic nerves.23.44 Thus, there seems to induces reflex voiding (the Bors Ice Water Test). 6 This be a considerable reorganization of reflex connections reflex does not occur in normal patients. Recently, it in the spinal cord following the interruption of descend has been shown in the cat that C-fiber bladder afferents ing pathways from the brain. C-fiber afferent-evoked are responsible for cold-induced bladder reflexes.2o reflexes which are weak and occur in only 60% of cats Arterial pressor responses induced by bladder disten with an intact neuraxis44 are facilitated; whereas AD sion occur in spinal injured patients and animals. 4,5,54 In afferent-evoked reflexes are completely eliminated in normal and spinal injured rats the increase in blood chronic spinal animals. These data indicate that two pressure induced by isometric bladder contractions or distinct central pathways (supraspinal and spinal) utiliz distension is suppressed by capsaicin treatment.22,54 ing different peripheral afferent limbs (A and C fiber) This suggests that C-fiber afferents mediate bladder can mediate detrusor to detrusor reflexes in the cat vascular reflexes in this species. Similar afferents may (Figure 6). The properties of the peripheral C-fiber be responsible for autonomic dysreflexia, in tetraplegic afferent receptors also appear to be changed in the patients. spinal injured cat. As mentioned above, C-fiber blad Capsaicin has also been evaluated clinically for the der afferents in the cat usually do not respond to treatment of various types of neurogenic disorders of Spinal injury and micturition we de Groat

501 the lower urinary tract. When administered intravesic peripheral organs. For example, vasoactive intestinal ally in concentrations between 100 uM and 2 mM polypeptide (VIP) immunoreactivity which is a marker capsaicin increases bladder capacity and reduces irritat for C-fiber afferents18,19,23,6o,61 and for a proportion of ive slmptoms in patients with hypersensitive blad the bladder afferent terminals in the sacral spinal cord ders1 ,55 and increases bladder capacity and reduces the of the cat18,23 is distributed over a wider area of the frequency of incontinence in patients with multiple lateral dorsal horn in paraplegic cats.62 This is consist sclerosis.56,57 The effects of high concentrations of ent with the concept of afferent axonal sprouting capasaicin in multiple sclerosis patients persisted for following spinal cord injury. In addition, the effects of weeks to months after treatment. intrathecal administration of VIP on bladder function In preliminary studies on a small population of spinal are changed in paraplegic cats. In normal cats VIP cord injured patients with detrusor hyperreflexia and inhibits bladder reflexes; whereas in paraplegic cats autonomic dysreflexia, intravesical capsaicin treatment small doses of VIP facilitate reflexes (Figure 9).23 increased bladder capacity, depressed the micturition These findings suggest that the action of a putative contraction pressure, decreased autonomic dysreflexia, C-fiber afferent transmitter is changed in concert with decreased urge incontinence and blocked the bladder the emergence of C-fiber reflexes. cooling reflex.58,59 These observations suggest that Although bladder reflexes recover in paraplegic capsaicin-sensitive, C-fiber bladder afferents are in patients the bladder does not empty efficiently due volved in several pathological conditions associated to uninhibited sphincter contractions (ie bladder with neurogenic bladder hyperactivity. sphincter dyssynergia).4-6 As shown in Figure 4C when the bladder contracts reflexly in the paraplegic patient Mechanisms of reflex plasticity following spinal cord the sphincter also contracts producing a functional injury outlet obstruction and a large volume of residual urine. The recovery of automatic micturition and C-fiber Thus viscerosomatic coordination in the lower urinary afferent-evoked bladder reflexes in paraplegic cats may tract appears to depend on control mechanisms located be dependent on multiple mechanisms: (1) elimination in the brain. of bulbospinal inhibitory pathways, (2) strengthening The failure of bladder sphincter coordination may of existing synapses or formation of new synaptic also lead to changes in spinal reflex pathways. It has connections due to axonal sprouting in the spinal cord, been speculated23,63 that bladder-sphincter dyssynergia (3) changes in synthesis, release, or actions of neuro and outlet obstruction in paraplegic animals could alter transmitters, (4) alterations in afferent input from the properties of bladder afferent pathways and reflex

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o 5 10 15 20 Minutes Effects of intrathecal administration of VIP on rhythmic bladder activity. In normal cats, large doses of VIP (1 to 10Figure ttg) inhibit9 bladder activity (A and B), whereas in chronic spinal cats smaller doses of VIP (0.1 to 1 ttg) facilitate bladder activity (C). Reproduced from de Groat et 199023 with permission we ai, Spinal injury and micturition We de Groat

502 mechanisms. This has been demonstrated in chronic diversion procedure to prevent bladder distension and spinal rats 4-6 weeks after transection of the spinal hypertrophy in the paraplegic animals.52,64 This pre cord.64 In these animals reflex voiding had developed; vented the afferent neuron hypertrophy suggesting that however voiding was inefficient (30-fold increase in factors released in the hypertrophied bladder were residual volume) due to tonic activity of the urethral responsible for the neural changes. A second approach sphincter.24,51 The bladder was markedly enlarged evaluated the neural changes induced by outlet obstruc (5-fold increase in weight) (Figure 10) and showed tion due to partial urethral ligation.26,65,66 Urethral prominent uninhibited contractions during cystometro ligation increased bladder work and induced bladder grams. Systemic capsaicin administration eliminated hypertrophy but did not eliminate the normal supra the uninhibited contractions but did not block the spinal control of the voiding.65 Outlet obstruction of micturition reflex.24 Anatomical tracing studies re 4-6 weeks duration induced hypertrophy of bladder vealed that the afferent neurons innervating the blad afferent (45% increase in size) and efferent neurons der were markedly increased in size (approximately (100% increase).26,66 This was accompanied by in 50% increase) (Figure 10) in the chronic spinal animals creased levels of nerve growth factor (NGF) in the compared to neurons in normal control animals.64 The bladder ,67 as well as an expansion of the bladder increase in size of afferent neurons could be associated afferent terminals in the cord and facilitation of the with sprouting of afferent axons in the bladder to spinal micturition reflex.26,65 Autoimmunization of rats innervate the larger target tissue. A larger cell body against NGF reduced the hypertrophy of efferent might be necessary to support a larger afferent terminal neurons indicating that changes in the levels of trophic field (Figure 10). This morphological change in bladder factors in the bladder can have a significant influence afferent neurons is consistent with the hypothesis on the neurons innervating the bladder. 67 Thus it is presented above that plasticity in the afferent limb of possible that spinal reflex mechanisms are affected the micturition reflex might contribute to the recovery indirectly by changes in organ functions which occur of voiding function in paraplegic animals. following spinal injury (Figures 11, 12). The signal for afferent neuron plasticity after spinal Electrical recordings from bladder afferent neurons injury has been explored using several experimental have revealed that the physiological properties of the approaches. One approach was to perform a urinary

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Functional obstruction EU:&Diagram � indicating the possible role of neuro Figuretrophic 11factors (NTF) in the neuroplasticity in afferent and efferent pathways to the urinary bladder of the rat follow ing spinal cord injury or partial urethral obstruction. In creased urethral resistance induced by functional urethral obstruction caused by enhanced external urethral sphincter (EUS) activity in paraplegic animals or by physical obstruc tion due to partial urethral ligation induces bladder hyper trophy and increased levels of NTF in the bladder. In the Hypertrophy physically obstructed animals NTF taken up by afferent and efferent terminals (dashed arrows) induces hypertrophy of bladder neurons in the major pelvic ganglion (MPG) and dorsal root ganglia (DRG) and increases the density of Changes in the innervation of the urinary bladder WGA-HRP labelled bladder afferent terminals in certain Figureof the 10rat in response to bladder hypertrophy. When the regions of the spinal cord. On the other hand, functional bladder smooth muscle mass increases following urethral obstruction induces a selective increase in bladder afferent obstruction, the neurons innervating the bladder also in neuron size and changes the electrophysiological properties crease in size. This has been detected as an increase in the of these neurons, but does not induce hypertrophy of cross sectional area of perikarya in sensory and autonomic efferent neurons in the MPG. This raises the possibility that ganglia. It also seems likely that axons in the bladder will neural activity or trophic substances (?) released by pre sprout to maintain a normal density of innervation in the ganglionic pathways from the spinal cord to the MPG hypertrophied organ negates the influence of peripheral neurotrophic factors Spinal injury and micturition WC de Groat

503 the brain and spinal cord. This control system performs Injury like a simple switching circuit to maintain a reciprocal relationship between the reservoir (bladder) and outlet ....--/ components (urethra and urethral sphincter) of the urinary tract. Spinal cord injury disrupts voluntary control and the normal reflex pathways that coordinate ".------. I Spinal cord bladder and sphincter function. Studies in animals indicate that recovery of bladder function following Changes � spinal injury is dependent upon the reorganization of In � organ function reflex pathways in both the peripheral and central nervous system. Part of this reorganization may be Diagram indicating an indirect mechanism that influenced by neural-target organ interactions (Figure Figuremight contribute12 to the changes in reflex circuitry in seg 12) that are mediated by neurotrophic factors released ments of spinal cord caudal to the level of spinal injury. by the peripheral organs. Interruption of descending pathways from the brain to the spinal cord can lead to changes in autonomic and somatic motor outflow to effector organs and thereby alter organ function. Experimental studies in spinal injured animals References indicate that changes in lower urinary function can alter the morphological and physiological properties of afferent de Groat WC, Booth AM, Yoshimura N. Neurophysiology of micturition and its modification in animal models of human neurons innervating the urinary bladder. It is speculated disease. In: Maggi CA (ed). The Autonomic Nervous System , that changes in afferent input to the spinal cord in paraple Vol. 3, Nervous Control of the Urogenital System . 1st edn. gic animals may have a significant influence on the reorgan Harwood Academic Publishers: London; 1993, pp 227-290. ization of spinal reflex circuitry that occurs following spinal 2 de Groat WC, Booth AM. Synaptic transmission in pelvic cord injury ganglia. In: Maggi CA (ed). 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A large Micturition, its Neurological Disorders and Sequelae . University majority (80%) of bladder afferent neurons from Park Press: Baltimore, 1971. 7 de Groat WC, Steers WD. Autonomic regulation of the urinary normal animals exhibited high threshold Na+ currents bladder and sex organs. In: Loewy AD, Spyer KM, (cds). that were resistant to tetrodotoxin (TTX) and low Central Regulation of Autonomic Functions . 1st edn. Oxford threshold K+ currents (A-type) that were activated University Press: Oxford, 1990, pp 310-333. at voltages between 60 and 50 mV. Due to this 8 de Groat WC. Neural control of urinary bladder and sexual combination of ion channels- the- neurons were rela organs. In: Bannister R, Mathias CJ, (eds). Autonomic Failure . 3rd edn. Oxford University Press: Oxford, 1992; pp 129-159. tively inexcitable. However in chronic spinal animals 9 Andersson K-E. Pharmacology of lower urinary tract smooth with bladder hypertrophy the majority (75%) of affer muscles and penile erectile tissues. Pharmacal Rev 1993; 45: ent neurons exhibited low threshold ITX-sensitive 253-308. Na+ currents and an absence of A-type K+ currents. 10 Somogyi GT, de Groat WC. Evidence of inhibitory nicotinic and facilitatory muscarinic receptors in cholinergic nerve ter These alterations in ion channels increased the excita minals of the rat urinary bladder. J Auton Nerv Syst 1992; bility of the afferent neurons. Similar alterations in 89-97. 37: electrical excitability at afferent receptors in the blad 11 Somogyi GT, Tanowitz M, de Groat We. M-1 muscarinic der could account for the apparent increased sensitivity receptor mediated facilitation of acetylcholine release in the rat of silent C-fiber afferents to bladder distension in urinary bladder but not in the heart. J Physiol (Land) 1994; 480: 81-89. paraplegic cats. 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Activation of unmyelin The lower urinary tract has two main functions: storage ated afferent fibres by mechanical stimuli and inflammation of the urinary bladder in the cat. J Physiol (Land) 1990; and periodic elimination of urine. These functions are 545-562. 425: regulated by a complex neural control system located in 16 Jiinig W, Koltzenburg M. Pain arising from the urogenital tract. Spinal injury and micturition we de Groat

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60 Kawatani M, Erdman SL, de Groat We. VIP and substance P neurons. J Auton Nerv Syst (in press) . in primary afferent pathways to the sacral spinal cord of the cat . 65 Steers WD, de Groat WC . Effect of bladder outlet obstruction J Comp Neurol 1985; 327-347, on micturition reflex pathways in the rat . J Urol 1988; 61 Kawatani M, Nagel 24J,1: de Groat We. Identification of 864-871. 140: neuropeptides in pelvic and pudendal nerve afferent pathways 66 Steers WD , Ciambotti J, Erdman S, de Groat WC . Morpho to the sacral spinal cord of the cat . J Comp Neurol 1986; logical plasticity in efferent pathways to the urinary bladder of 117-132, 249: the rat following urethral obstruction. J Neurosci 1990; 62 Thor K, Kawatani M, de Groat WC. Plasticity in the reflex 1943-1951 . 10: pathways to the lower urinary tract of the cat during postnatal 67 Steers WD et al. Nerve growth factor in the urinary bladder of development and following spinal cord injury. In: Goldberger the adult regulates neuronal form and function. J Clin Invest ME , Gorio A, Murray M, (eds) . Development and Plasticity of 1991; 1709-1715. the Mammalian Spinal Cord. Fidia Research Series Vol . III. 1st 68 Yoshimura88 : N, de Groat We. Changes in electrophysiological edn. Liviana Press: Padova, 1986, pp 65-80. and pharmacological properties of rat bladder afferent neurons 63 de Groat WC and Kruse MN. Central processing and morpho following spinal cord injury. J Urol 1995; 340A. logical plasticity in lumbosacral afferent pathways from the 69 Yoshimura N, Yoshida de Groat WC149:. Regional differences lower urinary tract . In: Mayer EA, Raybould HE, (eds) . Basic in plasticity of membrane0, properties of rat bladder afferent and Clinical Aspects of Chronic Abdominal Pain. Pain Research neurons following spinal cord injury. J Urol 1995; 262A. and Clinical Management. Elsevier Science Publishers: Amster 70 Yoshimura N, de Groat WC. Increases of neurofilament153: im dam, 1993; pp 219-235 . munoreactivity and reduction of capsaicin-sensitivity in rat blad Kruse MN, Bray LA, de Groat WC. Influence of spinal cord der afferent neurons following spinal cord injury. J Urol 1994; 64 injury on the morphology of bladder afferent and efferent 455A. 151: